This project, as its R21 Phase I milestone, will deliver a combination of conical nanopores having read length dimensions slightly less than 1 nm, and nucleobase-modified DNA oligonucleotides, where the passage of the DNA through the nanopore proceeds with a time constant of 10-100 microseconds per nucleotide, and where the ion current through the nanopore, during the time when the DNA is in transit, varies detectably depending on the nucleotide that is in the pore at the time that the current is measured. This nanopore-modified DNA combination will form the core of an extremely inexpensive technology to generate long reads of DNA sequence at the single molecule level. The research will exploit a decade of experience in the Martin laboratory preparing nanopores and engineering their chemical context, and an equal experience in the Benner laboratory working with nucleic acid analogs, polymerases that accept them, and practical applications of the combination. As specific aims, we shall: (a) prepare the nanotubes; (b) attach chemical functionality to the nanotubes; (c) prepare nucleoside triphosphates carrying different sized polyether dendrimers attached at the 5-position (for pyrimidines) and the 7-position (for 7-deazapurines); (d) use these triphosphates to synthesize modified DNA molecules. The nanopores will then be physically characterized to determine their ion transport dynamics, and in conjunction with the modified oligonucleotides, to find a combination that meets the R21 milestone specifications. If this milestone is passed, the R33 period will be used to develop sequence specific and randomly targeted primers that incorporate DNA, PNA, and tags that exploit an artificial genetic alphabet, and to develop improved processes for generating conical nanopores in a form suitable for large scale application. These will then be targeted against specific sequences extracted from mammalian genomes. [unreadable] [unreadable]